Cancer cells can utilize different invasion strategies to overcome physical arrest during confined migration through tissues with small pores. Cancer cell plasticity allows switches between different migration modes and transitions between single-cell and collective migration. The biophysical parameters that guide these decisions are poorly understood. In this work, we investigated the link between cell deformability and migration efficacy in constrictions of two mesenchymal cancer cell-types with similar invasion strategies: HT1080 fibrosarcoma cells and MV3 melanoma cells. To this end, we designed microfluidic platforms for (1) high-throughput cell deformability measurements and (2) migration through a variety of confining geometries. We measured different deformabilities for HT1080 and MV3 cells and correlated this with their migration efficacy through confinements. However, higher deformability and improved squeezing ability did not impact path selection at junctions of channels of different widths. Our findings show that cell deformability correlates with better squeezing abilities through confinements, but minimally impacts confinement directionality.

Tissue surface tension influences cell sorting and tissue fusion. Earlier mechanical studies suggest that multicellular spheroids actively reinforce their surface tension with applied force. Here we study this open question through high-throughput microfluidic micropipette aspiration measurements on cell spheroids to identify the role of force duration and spheroid deformability. In particular, we aspirate spheroid protrusions of mice fibroblast NIH3T3 and human embryonic HEK293T homogeneous cell spheroids into micron-sized capillaries for different pressures and monitor their viscoelastic creep behavior. We find that larger spheroid deformations lead to faster cellular retraction once the pressure is released, regardless of the applied force. Additionally, less deformable NIH3T3 cell spheroids with an increased expression level of alpha-smooth muscle actin, a cytoskeletal protein upregulating cellular contractility, also demonstrate slower cellular retraction after pressure release for smaller spheroid deformations. Moreover, HEK293T cell spheroids only display cellular retraction at larger pressures with larger spheroid deformations, despite an additional increase in viscosity at these larger pressures. These new insights demonstrate that spheroid viscoelasticity is deformation-dependent and challenge whether surface tension truly reinforces at larger aspiration pressures.

Cell spheroids are in vitro multicellular model systems that mimic the crowded micro-environment of biological tissues. Their mechanical characterization can provide valuable insights in how single-cell mechanics and cell-cell interactions control tissue mechanics and self-organization. However, most measurement techniques are limited to probing one spheroid at a time, require specialized equipment and are difficult to handle. Here, we developed a microfluidic chip that follows the concept of glass capillary micropipette aspiration in order to quantify the viscoelastic behavior of spheroids in an easy-to-handle, more high-throughput manner. Spheroids are loaded in parallel pockets via a gentle flow, after which spheroid tongues are aspirated into adjacent aspiration channels using hydrostatic pressure. After each experiment, the spheroids are easily removed from the chip by reversing the pressure and new spheroids can be injected. The presence of multiple pockets with a uniform aspiration pressure, combined with the ease to conduct successive experiments, allows for a high throughput of tens of spheroids per day. We demonstrate that the chip provides accurate deformation data when working at different aspiration pressures. Lastly, we measure the viscoelastic properties of spheroids made of different cell lines and show how these are consistent with previous studies using established experimental techniques. In summary, our chip provides a high-throughput way to measure the viscoelastic deformation behavior of cell spheroids, in order to mechanophenotype different tissue types and examine the link between cell-intrinsic properties and overall tissue behavior.

Spheroids are widely used in vitro 3D multicellular model systems that mimic complex physiological microenvironments of tissues. As different cell types vary in deformability and adhesion, the choice of (heterogeneous) cell composition will define overall spheroid mechanics, including their viscoelasticity and effective surface tension. These mechanical parameters directly influence cell sorting and possibly cell invasion into the extracellular matrix. Spheroid models therefore provide fundamental insights in the relation between cellular mechanics and important physiological processes, such as tissue formation, embryonic tissue remodeling, and cancer metastasis. In this review, we first summarize and compare current biophysical tools that probe mechanics of spheroids either from the outside or from within, then relate spheroid mechanics to cell mechanics and cell-cell adhesion, and subsequently discuss the role of spheroid mechanics alongside surrounding microenvironment parameters in (cancer) cell migration. We conclude by pointing out the research gaps and drawing the attention to novel techniques that could shed more light on the biophysical characterization of spheroids in the framework of tissue remodeling and cancer metastasis.